Direct Determination of ABO Blood Group Genotypes from Whole Blood Using PCR-Amplification of Specific Alleles Method
American Journal of BioScience
Volume 2, Issue 2, March 2014, Pages: 49-55
Received: Feb. 14, 2014;
Published: Mar. 30, 2014
Views 3257 Downloads 388
Kensaku Aki, Department of Cells and Immunity Analytics, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima, Japan
Kazuyoshi Kawazoe, Department of Pharmacy, Tokushima University Hospital, Tokushima, Japan
Azusa Izumi, Clinical Laboratory, Mie Prefectural Shima Hospital, Mie, Japan
Tomoki Tada, Department of Cells and Immunity Analytics, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima, Japan; Subdivision of Biomedical Laboratory Sciences, Graduate School of Health Sciences, The University of Tokushima, Tokushima, Japan
Kazuo Minakuchi, Department of Pharmacy, Tokushima University Hospital, Tokushima, Japan
Eiji Hosoi, Department of Cells and Immunity Analytics, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima, Japan
ABO antigens are known to be oligosaccharide antigens, and are widely expressed on the membranes of red blood cells and tissue cells. Therefore, the determining ABO blood group antigens is important in both transfusion and organ transplantation, and is one of the markers used for personal identification in forensics. The ABO blood group is currently determined by the presence of A and B antigens on red blood cells using serological tests in clinical laboratories. The gene sequences of the ABO blood group in chromosome 9q34.1-q34.2 have also been determined. Accordingly, it has become possible to genetically analyze the ABO blood group using molecular biological techniques. We recently developed an ABO genotyping method based on PCR amplification of specific alleles (PASA) using DNA extracted from blood and saliva. However, the extraction and purification of DNA is necessary prior to PCR because blood and saliva contain various substances that inhibit PCR. Furthermore, the PCR amplification of specific alleles (PASA) method requires specificity and stability for allele-specific amplification. Therefore, it is very difficult to use whole blood directly. Here, we described the development and use of ABO genotyping from whole blood using a commercially available reagent kit, which can effectively neutralize inhibitory substances present in the blood. In this study, for all genes of the six major ABO genotypes and cisA2B3 genotype of the AB variant, only specific bands were clearly amplified, whereas non-specific bands were not amplified at all. In addition, this method was able to determine ABO genotyping using 5-fold diluted fresh whole blood, or 5-fold diluted whole blood that was freeze-stored in 100 µL aliquots at -20 oC by subdivided for a maximum of 30 days. This analysis method to determine ABO blood group genotyping is simple and useful, and is expected to be used widely throughout research and clinical laboratories and forensic fields.
Direct Determination of ABO Blood Group Genotypes from Whole Blood Using PCR-Amplification of Specific Alleles Method, American Journal of BioScience.
Vol. 2, No. 2,
2014, pp. 49-55.
Landsteiner K (1900) Zur Kenntnis der antifermentativen, lytischen und agglutinierenden Wirkungen des Blutserums und der Lymphe. Zentralbl Bakteriol 27 : 357-362,
Lewis M, Anstee DJ, Bird GWG, Brodheim E, Cartron JP, et al. (1990) Blood group terminology 1990. The ISBT Working Party on Terminology for Red Cell Surface Antigens. Vox Sang 58 (2): 152-69.
Daniels GL, Fletcher A, Garratty G, Henry S, Jørgensen J, et al. (2004) Blood group terminology 2004: from the International Society of Blood Transfusion committee on terminology for red cell surface antigens. Vox Sang 87: 304-316.
Bernstein F (1924) Ergebnisse einer biostatistischen zusammenfassenden Betrachtung, ber die erblichen Blutstrukturen des Menschen. Klin Wochenschr 3: 1495-1497.
Shimada I, Kominato y, Hata N, Takizawa H (1999) DNA polymorphisms in the 5′-flanking sequence of human ABO blood group genes and their association with the alleles for the common ABO phenotypes. Legal Medicine 1: 217-225.
Svensson L, Hult AK, Stamps R, Ångström J, Teneberg S, et al. (2013) Forssman expression on human erythrocytes: biochemical and genetic evidence of a new histo-blood group system. Blood 121 (8): 1459-68.
Yamamoto F, Yamamoto M, Blancher A (2010) Generation of histo-blood group B transferase by replacing the N-acetyl-D-galactosamine recognition domain of human A transferase with the galactose-recognition domain of evolutionarily related murine alpha1,3-galactosyltransferase. Transfusion 50 (3): 622-30.
Saitou N, Yamamoto F (1997) Evolution of primate ABO blood group genes and their homologous genes. Mol Biol Evol 14 (4): 399-411.
Calafell F, Roubinet F, Ramirez-Soriano A, Saitou N, Bertranpetit J, et al. (2008) Evolutionary dynamics of the human ABO gene. Hum Genet 124 (2): 123-35.
Yamamoto F, McNeill PD, Hakomori S (1995) Genomic organization of human histo-blood group ABO genes. Glycobiology 5 (1): 51-58.
Yamamoto F, Hakomori S (1990) Sugar-nucleotide donor specificity of histo-blood group A and B transferases is based on amino acid substitution. J Biol Chem 265 (31): 19257-19262.
Bennett E P, Steffensen R, Clausen H, Weghuis DO, Geurts van kessel A (1995) Genomic cloning of the human hist-blood group ABO locus. Biochem.Biophys. Res. Commun 206 (1): 318-325.
Watkins W.M. (1980) Biochemistry and genetics of the ABO, Lewis, and P blood systems. Advances in Human Genetics (Harris H, Hirschhorn K, eds.). Plenum Press, New York. 1-136p.
Yamamoto F, Marken J, Tsuji T, White T, Clausen H, et al. (1990) Cloning and characterization of DNA complementary to human UDP-GalNAc: Fucα1→2Galα1→3GalNAc transferase (histo-blood group A transferase) mRNA. J. Biol. Chem. 265:1146-1151.
Yamamoto F, Clausen H, White T, Marken J, Hakomori S (1990) Molecular genetic basis of the histo-blood group ABO system. Nature 345: 229-233.
Hosoi E, Yoshimoto K (1993) Genetic analysis of the genotype of ABO and cisAB blood group. Jpn J Clin Pathol 41 (10): 1133-1140.
Hosoi E (1996) Direct determination of ABO and cisAB blood group genotypes using polymerase chain reaction amplification of specific alleles (PASA)-method. Jpn J Clin Pathol 44 (8): 783-790.
Hosoi E (1997) Genetic analysis of the ABO blood groups and application of the clinical laboratories. Jpn J Clin Pathol 45 (2): 148-156.
Hosoi E, Hirose M, Hamano S, Kuroda Y (1998) Detection of histo-blood group ABO mRNA in human chronic myeloid leukemia cell lines using reverse transcription-polymerase chain reaction (RT-PCR). Cancer Lett 133 (2): 191-196.
Ogasawara K, Yabe R, Uchikawa M, Saitou N, Bannai M, at al. (1996) Molecular genetic analysis of variant phenotypes of the ABO blood group system. Blood 88 (7): 2732-2737.
Ogasawara K, Yabe R, Uchikawa M, Bannai M, Nakata K, et al (1998) Different alleles cause an imbalance in A2 and A2B phenotypes of the ABO blood group. Vox Sang 74 (4): 242-247.
Aki K, Izumi A, Hosoi E (2012) The evaluation of histo-blood group ABO typing by flow cytometric and PCR-amplification of specific alleles analyses and their application in clinical laboratories. J Med Invest 59 (1-2): 143-51.
Panaccio M, Lew A (1991) PCR based diagnosis in the presence of 8% (v/v) blood. Nucleic Acids Res 19 (5):1151.
Mercier B, Gaucher C, Feugeas O, Mazurier C (1990) Direct PCR from whole blood, without DNA extraction. Nucleic Acids Res 18 (19): 5908.
McCusker J, Dawson MT, Noone D, Gannon F, Smith T (1992) Improved method for direct PCR amplification from whole blood. Nucleic Acids Res. 20 (24): 6747.
Casareale D, Pottathil R, Diaco R (1992) Improved blood sample processing for PCR. PCR Methods Appl 2 (2): 149-53.
Burckhardt J (1994) Amplification of DNA from whole blood. PCR Methods Appl 3 (4): 239-43.
Akane A, Matsubara K, Nakamura H, Takahashi S, Kimura K (1994) Identification of the heme compound copurified with deoxyribonucleic acid (DNA) from bloodstains, a major inhibitor of polymerase chain reaction (PCR) amplification J Forensic Sci, 39: 362–372.
Al-Soud WA, Jonsson LJ, Rådström P (2000) Identification and characterization of immunoglobulin G in blood as a major inhibitor of diagnostic PCR. Clin Microbiol, 38: 345–350.
Al-Soud WA, Rådström P (2001) Purification and characterization of PCR-inhibitory components in blood cells. J Clin Microbiol, 39: 485-493.
Satsangi J, Jewell DP, Welsh K, Bunce M, Bell JI (1994) Effect of heparin on polymerase chain reaction. Lancet, 343:1509–1510.
Nishimura N, Nakayama T, Tonoike H, Kojima K, Kato S (2000) Direct polymerase chain reaction from whole blood without DNA isolation. Ann Clin Biochem (Pt 5):674-80.
Nishimura N, Nakayama T, Tonoike H, Kojima K, Shirasaki Y, at al (2002) Various applications of direct PCR using blood samples. Clin Lab 48 (7-8): 377-84.
Hummei S, Schmidt D, Kahle M, Herrmann B (2002) ABO blood group genotyping of ancient DNA by PCR-RFLP. Int J Leqal Med 116 (6): 327-33.
Lee JC, Hsieh HM, Teng HF, Lo SC, Linacre A (2009) ABO genotyping by single strand conformation polymorphism--using CE. Electrophoresis 30 (14): 2544-8.
Fregel R, Maca-Meyer N, Cabrera VM, González AM, Larruga JM (2005) Description of a simple multiplex PCR-SSCP method for ABO genotyping and its application to the peopling of the Canary Islands. Immunogenetics 57 (8): 572-8.
Sung Ho L, Geon P, Young Geun Y, Seung Gwan L, Suhng Wook K (2009) Rapid ABO Genotyping Using Whole Blood without DNA Purification. Korean J Lab Med 29 (3): 231-7.
Hwan Young L, Myung Jin P, Na Young K, Woo Ick Y, Kyong-Jin S (2011) Rapid Direct PCR for ABO Blood Typing. J Forensic Sci 56: 179-82.